FIG. 1 shows the general formula (I) showing the molecular structure of conjugated ursodeoxycholic acids, the final products of a method in accordance with the present invention. Referring to the Figure, a conjugated ursodeoxycholic acid is a bile acid wherein hydroxyl groups are located at 3.alpha. and 7.beta. positions thereof and the carboxyl group located in a side chain thereof is conjugated with an amino acid (including artificially produced amino acids).
Out of the conjugated ursodeoxycholic acids, tauroursodeoxycholic acid, a ursodeoxycholic acid conjugated with taurine, is the major substance of the gall-bladder of a bear. This compound is a potential raw material for producing a ursodeoxycholic acid which has recently been employed as a cholagogue or a cholesterol solubilizer. In other words, a ursodeoxycholic acid can be produced by deconjugation of tauroursodeoxycholic acid. Since the supply of good gall-bladders of bears is fairly limited, it has been difficult to satisfy the entire quantity of medical demands with the aforementioned natural supply. This is the reason why chemical synthesis processes have been employed for producing ursodeoxycholic acid.
However, each of the chemical synthesis processes available in the prior art comprises plural complicated steps. Since the number of steps each of which is fairly complex, is sometimes as large as 7, they are involved with drawbacks wherein the process time is long, the process is complicated, the efficiency is inferior due to the purification processes each of which is needed after each individual step, and the yield is unsatisfactory due to side reactions each of which inevitably occurs accompanying each individual step.
Aiming at removal of these drawbacks, various one step methods have been proposed for producing ursodeoxycholic acid by means of microbial transformation wherein a .beta.-hydroxyl group is introduced to be bonded with a carbon atom which is located at 7-position of the steroid nucleus (ring) of lithocholic acid.
We were successful in the development of a one step method for producing free ursodeoxycholic acid from free lithocholic acid, which comprises subjecting lithocholic acid, the starting material, to the action of any of the specific microorganisms or moulds belonging to Fusarium.
However, the transformational reaction substrate for this one step method for producing ursodeoxycholic acid is free lithocholic acid which does not necessarily readily dissolve in a cultivation medium and/or a reaction medium. Accordingly, this hydrophobic nature of the tranformational reaction substrate readily causes a low degree of microbial transformation in the aforementioned method, thus causing a low of yield, a slow reaction rate and a low degree of productivity.
In view of the fact that free ursodeoxycholic acid is converted, in the human body, to conjugated ursodeoxycholic acids, when it is internally applied, we assumed that some conjugated ursodeoxycholic acids would have medicinal value which is equivalent or superior to that of free ursodeoxycholic acid. Based on this assumption, and also based on the fact that conjugated lithocholic acids readily dissolve in water, we considered that a method for producing one or more conjugated ursodeoxycholic acids by means of microbial transformation which comprises subjecting one or more of the conjugated lithocholic acids to the action of one or more of the microorganisms which have capability for producing conjugated ursodeoxycholic acids, would remove the aforementioned drawbacks.